Visual DNA Microarray Coupled with Multiplex-PCR for the Rapid Detection of Twelve Genetically Modified Maize

다국어 초록 (Multilingual Abstract)

We herein developed a visual DNA microarray system coupled with multiplex PCR (m-PCR) to rapidly detect twelve genetically modified maize (GMM). The microarray comprised short oligonucleotide probes complimentary to the specific gene region for twelve different GMM. The m-PCR products annealed to the microarray probe were reacted with streptavidin-alkaline phosphatase conjugate and nitro blue tetrazolium/5-bromo-4-chloro-3ʹ-indolylphosphate, p-toluidine salt (NBT/BCIP), resulting in blue spots that are easily visualized by unaided eyes for qualitative analysis. To ensure the reliability of this method, positive and negative hybridization controls were used in DNA microarray. Commercial GM materials (GMM: Bt176, Bt11, MON810, GA21, T25,MON88017, NK603, MON863, MON89034, DAS-59122-7, TC1507, MIR604; GM cotton: (MON1445, MON15985); GM soybean (Monsanto Roundup Ready soybean 40-3-2)) and non-GM materials were identified by this method and further confirmed by PCR and sequencing. The results showed that each probe consistently identified its corresponding GMM target very quickly and in a cost-effective and more time efficient way. The limit of detection is 0.5% for Bt176, Bt11, T25, MON88017, DAS59122-7, MON89034 and 1% for MON810, MIR604, GA21, MON863, NK603, TC1507. This method is advantageous because of rapid detection, cost-effectiveness and ease of use. These high specificity and sensitivity results demonstrate the feasibility of using this method in routine analysis of GMOs.

다국어 초록 (Multilingual Abstract)

We herein developed a visual DNA microarray system coupled with multiplex PCR (m-PCR) to rapidly detect twelve genetically modified maize (GMM). The microarray comprised short oligonucleotide probes complimentary to the specific gene region for twelve different GMM. The m-PCR products annealed to the microarray probe were reacted with streptavidin-alkaline phosphatase conjugate and nitro blue tetrazolium/5-bromo-4-chloro-3ʹ-indolylphosphate, p-toluidine salt (NBT/BCIP), resulting in blue spots that are easily visualized by unaided eyes for qualitative analysis. To ensure the reliability of this method, positive and negative hybridization controls were used in DNA microarray. Commercial GM materials (GMM: Bt176, Bt11, MON810, GA21, T25, MON88017, NK603, MON863, MON89034, DAS- 59122-7, TC1507, MIR604; GM cotton: (MON1445, MON15985); GM soybean (Monsanto Roundup Ready soybean 40-3-2)) and non-GM materials were identified by this method and further confirmed by PCR and sequencing. The results showed that each probe consistently identified its corresponding GMM target very quickly and in a cost-effective and more time efficient way. The limit of detection is 0.5% for Bt176, Bt11, T25, MON88017, DAS59122-7, MON89034 and 1% for MON810, MIR604, GA21, MON863, NK603, TC1507. This method is advantageous because of rapid detection, cost-effectiveness and ease of use. These high specificity and sensitivity results demonstrate the feasibility of using this method in routine analysis of GMOs.

참고문헌 (Reference)

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